Stanični odgovor bakterije Escherichia coli na pogreške u biosintezi proteina na izoleucinskim položajima

doctoral thesis

• Title (english): Cellular response to proteome-wide isoleucine mistranslation in Escherichia coli
• Author: Marija Pranjić
• Mentor: Marko Močibob (mentor)
• Committee member: Ivana Ivančić Baće (predsjednik povjerenstva)
• Committee member: Anita Horvatić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Chemistry); Zagreb
• Defense date and country: 2024-05-20, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Chemistry
• Universal decimal classification (UDC): 54 - Chemistry. Crystallography. Mineralogy

Abstract

Istražen je stanični odgovor bakterije Escherichia coli na pogreške u biosintezi proteina na izoleucinskim položajima (mistranslacija) metodama klasične mikrobiologije i kvantitativne proteomike uz uporabu spektrometra masa visoke rezolucije. Uspoređena je toksičnost kanonske i nekanonske mistranslacije, odnosno zamjene izoleucina proteinogenim valinom i zamjene izoleucina neproteinogenim norvalinom. Pokazano je da Escherichia coli može podnijeti vrlo visoku razinu mistranslacije (do 20 %). Rast u nutritivno siromašnim uvjetima uz dodatak norvalina ili valina uzrokuje značajnu mistranslaciju na izoleucinskim pozicijama u divljem tipu bakterije. Mistranslacija inhibira stanični rast, smanjuje otpornost stanice na toplinski stres te izaziva SOS-odgovor i filamentaciju stanica u stacionarnoj fazi rasta. Izoleucinska mistranslacija pojačava ekspresiju glavnih proteaza i šaperonskih sustava. Uslijed mistranslacije mijenjaju se i interaktori glavnog bakterijskog šaperona DnaK koji su ukazali na preferentnu destabilizaciju proteina uključenih u proces translacije u stanici. Nekanonska mistranslacija mijenja ekspresiju većeg broja proteina i izaziva veći inhibitorni učinak na rast stanica od kanonske mistranslacije.

Abstract (english)

he cellular response of Escherichia coli to errors in protein biosynthesis at isoleucine positions (mistranslation) was investigated using classical microbiological methods and mass-spectrometry-based quantitative proteomics. The toxicity of canonical and non-canonical mistranslation was compared, e.g. the substitution of isoleucine with proteinogenic valine and the substitution of isoleucine with non-proteinogenic norvaline. Escherichia coli can tolerate very high levels of isoleucine mistranslation (up to 20%). Growth in nutritionally poor conditions with the addition of norvaline or valine causes significant isoleucine mistranslation in wild-type bacteria. Mistranslation inhibits cellular growth, reduces the cell's resilience to heat stress and causes the SOS response and filamentation in the stationary phase. Isoleucine mistranslation causes upregulation of major proteases and chaperone systems. The interactors of the main bacterial chaperone DnaK change during mistranslating conditions indicating preferential destabilization of proteins involved in translational processes. Non-canonical mistranslation causes higher proteome dysregulation and a higher inhibitory effect on cellular growth than canonical mistranslation.

• Keywords: izoleucil-tRNA-sintetaza; mistranslacija; norvalin; spektrometrija masa; valin
• Keywords (english): isoleucyl-tRNA synthetase; mass spectrometry; mistranslation; norvaline; valine
• Language: croatian
• URN:NBN: urn:nbn:hr:217:464649
• Promotion: 2024
• Project: Number: IP-2016-06-6272; Title: Aminoacil-tRNA-sintetaze kao čuvari standardnog genetičkog koda; Title: Aminoacyl-tRNA synthetases as gatekeepers of the standard genetic code; Acronym: AARSCODE; Leader: Ita Gruić-Sovulj; Jurisdiction: Croatia; Funder: Hrvatska zaklada za znanost; Funding stream: Research Projects
• Study programme: Title: Doctoral study; Study programme type: university; Study level: postgraduate; Academic / professional title: doktor/doktorica u području prirodnih znanosti (doktor/doktorica u području prirodnih znanosti)
• Type of resource: Text
• Extent: 123, XIX str. ; 30 cm
• File origin: Born digital
• Access conditions: Open access
• Created on: 2024-05-22 12:33:23